This study aimed to investigate the physiological differences, physical characteristics, and some plasma hormones in chicks hatched from multi-stage and single-stage incubator systems. A total of 7200 eggs were obtained from broiler breeder flock (cobb-500) at 37 weeks of age. Eggs were randomly divided into two groups, each of 3600 eggs, with three replicates of 1200 eggs. The first group was multi stage incubator and the second group was single stage incubator. Results showed that there was no significant difference between the two incubation systems in terms of hatchability. Furthermore, hatchability percent for fertile egg have significant differences favor a single-stage machine, this demonstrates the superiority of single stage machines. There was a significant difference between the two systems for the percentage of dead pipped embryonic and late embryonic with the single stage system having a lower percentage of both. Chicks hatched from the single stage incubator had a heavier body weight and longer body, leg, and gut. The blood plasma of chicks that were hatched using a single stage system contained a higher concentration of both total proteins and globulins, it was also observed the activity of liver enzymes especially AST and increasedT4 concentration compared to blood plasma of chicks hatched multi stage system. The result reflected an increase in the value of triiodothyronine, triiodothyronine to thyroxine ratio and corticosterone concentration (0.34 nmol/l, 0.02 ratio, and 18.5 ng/ml, respectively) in the blood plasma of the hatched chicks in multi-stage hatching machines.